Methods of patterning a monolayer

ABSTRACT

A method of patterning a monolayer including the steps of providing a monolayer of a compound on a substrate, positioning a near field light source in relation to the monolayer so that light from the light source irradiates the monolayer in the near field regime, the wavelength of the light being suitable to react with molecules in the monolayer and initiate a photochemical reaction, and patterning the monolayer by causing a relative movement of the monolayer and the near field light source, the relative movement corresponding to a desired pattern.

This invention relates to methods of patterning a monolayer, and tomethods of selectively coupling a molecular species to such patternedmonolayers.

In the past ten years there has been a great deal of scientific interestin the fabrication of micropatterned organic materials. This interest isdriven by the potential utility of such materials in a wide range oftechnological applications. These include the development of hybridorganic-metallic or organic-semiconductor electronic devices (including,for example, molecular electronics and lab-on-a-chip technology) and thepreparation of biological arrays. Array based systems are importanttools for research in the biological sciences. The applications arewide-ranging, and include high-throughput DNA sequencing, biomolecularanalysis and combinatorial testing methods. In many of theseapplications, high density arrays are utilised (for example, chipsdesigned for DNA sequencing often incorporate thousands of individuallyaddressable locations to each of which a distinct oligonucleotidesequence is attached). Demands for increasing analytical capability arecreating pressure to generate ever more dense arrays, necessitating theattainment of decreasing feature sizes. An additional benefit ofdecreasing feature size is increased sensitivity. The ultimatesensitivity in biological analysis would be represented by the detectionand analysis of single molecules. Array based methods offer the promiseof ultra-high sensitive analysis, provided that methods are developedfor creating adequately small features. There is consequently widespreadinterest in methods for the fabrication of biological structures andarrays with nanometre dimensions, although few successful methodscurrently exist. Outside of the biological sciences, there is muchinterest in the miniaturisation of electronic circuitry to facilitatethe fabrication of microprocessors and information storage systems withgreater capability. There is also much interest in the development ofmethods for the fabrication of miniaturised electronic devices from newmaterials, where greater ease of fabrication is possible, or where newmaterials (for example, organic materials) might be used in place ofconventional ones. All of these technologies place a requirement for newmethods for the fabrication of nanometre scale structures and for thecontrol of molecular structure at the nanometre scale. Whilst a numberof methods are available for creating micron-scale patterns, there arevery few that are capable of being used to fabricate nanostructuredmaterials (ie materials with feature sizes less than 100 nm).

A variety of types of materials have been patterned, including polymersand organic monolayers. Self-assembled monolayers (SAMs) have emerged aspotentially very useful materials in such applications. These materialsare formed by the adsorption of alkanethiols HS(CH₂)_(n)X onto gold,silver and some other surfaces. The thiol-on-gold SAM is the most widelystudied system. The main components (see FIG. 1) are a thiol group 10,which tethers the adsorbate to a substrate such as gold through a strongspecific interaction (effectively a covalent bond), a tail group X 14which is directed away from the surface, and an alkyl chain 16 thatlinks the two together. The properties of the surface may be controlledby changing the structure of the adsorbate molecule. For example, thewettability may be controlled by changing the tail group. Methylterminated SAMs are very hydrophobic—water contact angles are in therange 99-115°, depending on the length of the alkyl chain—while SAMswith polar tail groups (eg OH, COOH) may be hydrophilic, with contactangles typically less than 15°.

The most widely used method for making patterned SAMs is microcontactprinting (μCP). This simple, flexible method involves creating a siliconrelief mask by photolithography, and casting silicone elastomer onto themask. After curing, the silicone may be removed from the master and therelief features inked with a solution of a thiol, which may then betransferred to a gold substrate by stamping. This method is very easy touse and versatile. There is considerable interest in extending itscapability to the nm scale. However, it is difficult to accuratelycreate features with adequately well-defined dimensions at the nm scale.The best results obtained thus far have been by squeezing a stamp withmicron scale features. The resulting compression results in features assmall as 100 nm, but there are clear limitations in the types offeatures that may be created using the method.

An alternative approach is disclosed in U.S. Pat. No. 5,514,501. Thisphotolithographic procedure is depicted in FIG. 2, and involved placinga mask 20 over the surface of a thiol 22 of one particular chemistry (inFIG. 2, this is a carboxylic acid terminated thiol). The sample is thenexposed to UV light 24 (FIG. 2(a)). In exposed areas, the thiols areconverted to alkylsulphonates, RSO₃26 (FIG. 2(b)). While the thiols arebound strongly to the underlying gold, the sulphonates are only weaklybound. The sample is then dipped into a solution of a second thiol,displacing the sulphonates and adsorbing the second thiol 28 at thesurface (FIG. 2(c)). If this second thiol 28 has a different tail groupchemistry (eg, methyl as shown in FIG. 2), the result will be a chemicalpattern, in which the masked areas contain the original chemistry andthe exposed areas contain a new chemistry. This method is very effectiveand easy to use. The resulting patterns have clean, well-definedchemical structures and good edge-definition. The drawback is that itrelies upon exposure through a mask, which imposes a limitation onfeature sizes: conventional photolithography using UV light is notcapable of creating nanostructures because diffraction occurs when thefeatures in the mask become smaller than half the wavelength of thelight used.

Nanoscale patterned SAMs have been produced using a technique known asdip-pen nanolithography (DPN). This method, which is not aphotolithographic method, involves dipping the tip of an atomic forcemicroscope in a solution of an alkane thiol, and then using the tip totransfer the thiol to a gold surface much as a pen would write with inkon a sheet of paper. DPN as been successfully used to create featureswith dimensions of a few tens of nanometres.

Scanning near field optical microscopes (SNOMs) have been in usage sincethe early 1990s. In a SNOM, a narrow optical fibre (having an internaldiameter as small as 50 nm) is brought in close proximity to a samplesurface. Under such conditions, as a result of a near field effect,light may be transmitted through the aperture without undergoingdiffraction, even though the fibre aperture is smaller than half thewavelength of the light. SNOMs have been used principally as opticalprobes for surface characterisation. Some attempts have been made to useSNOMs in order to perform very high resolution photolithography. Inparticular, attempts have been made to pattern conventional photo-resistmaterials using a SNOM. However, disappointing results have beenobtained, and it is believed that the reason for this lack of success isthat either light from the SNOM tends to diverge within the resist layerand/or that thermal transfer (ie, heating) is occurring, with theconsequence that the feature sizes have generally been larger than 100nm. A small number of investigations have reported feature sizesslightly smaller than 100 nm, but the features have rarely been sharplydefined. International Publication WO98/58293 discloses a number oftechniques for immobilising macromolecules on a surface, one of which isinitiated by the use of optical near field microscopy to photoactivatefunctional groups on a layer which is disposed on a surface. Thetechnique appears to be rather speculative in nature: no experimentaldetails are provided (such as the depth of the layer), and noquantitative indication is provided of the feature size which may beproduced. The general leaning in the art is that SNOM techniques areunsuitable for high resolution nonstructural patterning.

The present invention overcomes the above mentioned problems,limitations and disadvantages, and provides a photolithographic methodwhich can enable high resolution patterning of substrates on a scale ofa few tens of nanometres.

According to a first aspect of the invention there is provided a methodof patterning a monolayer of a compound comprising steps of:

-   -   providing a monolayer on a substrate;    -   positioning a near field light source in relation to the        monolayer so that light from the light source irradiates the        monolayer in the near field regime, the wavelength of the light        being suitable to interact with molecules in the monolayer and        thereby initiate a photochemical reaction; and    -   patterning the monolayer by causing relative movement of the        monolayer and the near field light source, the relative movement        corresponding to a desired pattern.

In this way it is possible to fabricate structures having well definedchemistries and dimensions. A wide range of feature sizes can beproduced, including nanostructures. Feature sizes in the range of a fewtens of nanometres are possible. Surprisingly, it has been found thatnear field techniques can be used to produce such well defined features,despite the relatively poor spatial resolution achieved with prior arttechniques which utilised a SNOM to pattern photoresists. It is believedthat the improvement afforded by the present invention is, at least inpart, due to the provision of a monolayer on the substrate. It isbelieved that light from the near field light source does not divergethrough the monolayer and/or cause local heating effects, in contrast tothe conventional photoresists utilised in the prior art.

The monolayer may be a SAM, such as a thiolate SAM, or monolayers ofalkylsilanes, carboxylic acids or phosphonic acids. Langmuir-Blodgettfilms are possible alternatives.

Preferably, the near field light source comprises a scanning near fieldmicroscope (SNOM). It should be noted that some manufacturers term suchinstruments near field scanning optical microscopes (NSOMs), and thatfor the avoidance of doubt, such terms should be regarded as beingequivalent to SNOM. The use of other forms of near field light sourcesis within the scope of the invention.

The molecules in the monolayer which absorb light from the light sourcemay be converted by the photochemical reaction into a weakly boundspecies which is less strongly bound to the substrate than the moleculeswhich originally comprise the monolayer. The method may further comprisethe step of displacing the weakly bound species from the substrate witha displacing species. The displacing species may comprise a component ofa chemical etch. The chemical etch may etch the substrate.

The photochemical reaction may be a photooxidation reaction. Thephotooxidation reaction may be used to convert molecules in the SAM intoa weakly bound species in the manner described above. In such instances,it is likely that a ‘head’ group, in contact with the substrate, will beoxidised. However, other oxidative processes, such as photooxidation ofone ‘end’ group or of an alkyl chain, are possible. Alternatively, adifferent type of photochemical reaction may be initiated.Photoactivation of a group may be performed, for example to attachmolecules (such as biological molecules) to an end group, or to initiatecross linking of groups such as diacetylenic groups. In the latterinstance, the position of the diacetylenic groups may be the subject ofvariation; generally they are about half way along an alkyl chain. Thephotochemical reaction might comprise a unimolecular reaction or even ahalf-reaction. Furthermore, desorption or ablation may occur, either asa main patterning mechanism or in conjunction with a photochemicalreaction, and such processes should be considered to be a photochemicalreaction for the purpose of the present invention.

The molecules in the SAM may comprise thiolates. The SAM may be formedfrom thiols, which may comprise alkylthiols. The thiols may be of theformula HS(CH₂)_(n)X where X is an end group. The end group X may be anyfunctional group which provides desired characteristics, eg. reactivity,hydrophobicity, etc. The end group X may be selected from the groupconsisting of CH₃, CO₂H and OH. Typically n is in the range 0 to 20.Other possibilities are within the scope of the invention. For example,the thiolate may be partially or per fluorinated, and other end groups,such as NH₂, CF₃, halogen, etc, may be used. Other compounds capable ofproviding thiolate SAMs, such as dialkyl sulphides and dialkyldisulphides, might be used in place of thiols.

In the instance in which the molecules in the SAM comprise thiolates,and the photochemical reaction is a photooxidation reaction, thephotooxidation reaction may oxidise thiolate moieties adsorbed on thesubstrate to sulphonate moieties. The sulphates produced by such aphotooxidation are relatively weakly bound to the substrate. Thesulphonates (or any other weakly bound species) may be displaced fromthe substrate with a displacing species which forms a thiolate compoundon the substrate. Thiols, dialkyl sulphides and dialkyl disulphides arecandidates as displacing species. It should be noted that there iscontroversy in the literature over the mechanism of the photooxidationto sulphonates. Many studies suggest the primary mechanism isozonolysis, a mechanism which would be inconsistent with the mechanismof the present invention, in which close positioning of SNOM and SAM areeffected in order to achieve near field conditions and directinteraction of light from the SNOM with the SAM.

The substrate may comprise gold or silver. Other candidates includecopper, platinum, iridium, palladium, rhodium, mercury, osmium,ruthenium, and semiconducting materials such as gallium arsenide,iridium phosphide, mercury cadmium telluride and silicon.

The light source may provide near UV light. For example, in order tooxidise thiolate moieties to sulphonate moieties, wavelengths of around240 to 260 nm are desirable. The skilled reader will appreciate that theoptimal wavelength or wavelength range will depend on numerous featuressuch as the wavelength dependence of both the absorption coefficients ofthe molecular species in question and the quantum yield for the desiredphotochemical reaction, and the nature and availability of possiblelight sources (typically laser light sources). Other regions of theelectromagnetic spectrum might be used, depending on the photochemicalscheme employed. For example, vacuum UV light might be used to patternsilane monolayers, such as the 193 nm output of an ArF excimer laser.

The patterning may comprise features having at least one dimension in adirection parallel to the substrate which is less than 100 nm,preferably less than 50 nm.

According to a second aspect of the invention there is provided a methodof selectively coupling a molecular species to a surface, comprising thesteps of:

-   -   pattering a monolayer using a method in accordance with the        first aspect of the invention to provide one or more features;        and    -   coupling the molecular species to a compound present on the        substrate.

In this way it is possible to produce a vast range of devices havingstructures which exhibit a desired functionality, in which thedimensions associated with such structures are of the order of tens ofnanometres.

The molecular species may be coupled to the monolayer, ie, the couplingmay be to the monolayer itself, rather than to the regions which havebeen patterned.

Alternatively, the molecular species may be coupled to the features. Forexample, in the instance in which a displacing species has displaced aweakly bound species from the substrate in the patterned regioncorresponding to the features, the molecular species may be coupled tothe displacing species.

Alternatively still, an intermediate compound or compounds may becoupled to a portion of the patterned monolayer (which may be themonolayer or the features), and the molecular species coupled to theintermediate compound.

The coupling may comprise adsorption of the molecular species onto thecompound. Such coupling schemes are disclosed in U.S. Pat. No.5,514,501, the contents of which are incorporated by reference.

The coupling may comprise covalently bonding the molecular species tothe compound. Many such coupling schemes are known in the literature:for example, P. Wagner, P. Kerned, M. Hegner, E. Ungewickell and G.Semenza, FEBS letters 356 (1994); N. Patel, M. C. Davies, M. Hartshorne,R. J. Heaton, C. J.Roberts, S. J. B. Tendler and P. M. Williams,Langmuir 13 (1997) 6485-6490; G. J. Leggett, C. J. Roberts, P. M.Williams, M. C. Davies, D. E. Jackson and S. J. B. Tendler, Langmuir 9(1993), 2356-2362; and G. J. Leggett, C. J. Roberts, P. M. Williams, M.C. Davies, D. E. Jackson and S. J. B. Tendler, Langmuir, 9 (1993),2356-2362, and W. Knoll, L. Angermaier, G. Batz, T. Fritz, S. Fujisawa,T. Furano, H. J. Guder, M, Hara, M. Liley, K. Niki and J. Spinke, Synth.Met 61 (1993) 5, the contents of which are hereby incorporated byreference. Thus, for example, water soluble carbodimides either with orwithout the use of N-hydroxysuccinimide can be used to attach proteinsto carboxylic acid terminated SAMs, and photopatterned SAMs can bederivatised with perfluorinated molecules (see, for example, D. A. Huttand G. J. Leggett, Langmuir, 13 (1997) 2740-2748) the contents of whichare hereby incorporated by reference). Thiols terminated in biotin canbe absorbed onto the substrate—perhaps being used as the displacingspecies. The biotin terminated thiols can be used to bind streptavidinmolecules, which then bind biotinylated antibodies.

The molecular species may be a biological molecule. The biologicalmolecule may be a protein, a DNA strand, an RNA strand, anoligonucleotide or an enzyme. In this way, devices to perform operationssuch as biosensing, sequencing, synthesis and chemical or biologicalanalysis can be fabricated comprising, if necessary, nanostructures.

Embodiments of methods in accordance with the invention will now bedescribed with reference to the accompanying drawings, in which:

FIG. 1 shows a self assembled monolayer (SAM);

FIG. 2 shows a prior art patterning technique;

FIG. 3 shows a) the formation of a SAM on a substrate, b) treatment ofthe SAM with a SNOM, c) photooxidised regions of the SAM, and d), thedisplacement of oxidised molecules by a displacing species; and

FIG. 4 shows a lateral force microscopy image of a patterned hydroxylterminated SAM; and

FIG. 5 shows (a) a lateral force microscopy image of a patternedcarboxylic acid terminated SAM and (b) a close view of a line present in(a);

FIG. 6 shows (a) treatment of a SAM on a gold surface with a SNOM, (b)photooxidised regions of the SAM, and (c ) etching of the gold surface;and

FIG. 7 shows (a) an atomic force microscopy image of an etched SAM/goldsurface and (b) a cross sectional view through the image of (a).

FIG. 3 depicts a method of patterning a SAM in accordance with theinvention. In step (a) a SAM 30 is formed on a suitable substrate 32. Instep (b), a SNOM is disposed very close to the SAM 30 so thatinteraction in the near field regime can occur. The SNOM tip 34 is shownin FIG. 3(b), the tip 34 typically comprising a narrow optical fibre(eg, of internal diameter ca. 50 nm). The SNOM irradiates the SAM 30with light 36 of an appropriate wavelength to initiate a desiredphotochemistry. Under near field conditions, the light 36 passes throughthe aperture formed by the end of the optical fibre 34 withoutundergoing diffraction, even though the diameter of the aperture may beless than half of the wavelength of the light. The SNOM is moved inrelation to the SAM 30 in order to effect the desired pattern on the SAM30. FIG. 3(c) shows the result of the irradiation of the SAM 30 withlight 36 from the SNOM: highly localised conversion occurs of themolecules in the SAM 30 into a more weakly bound species 38. Whereverthe SNOM fibre has travelled, the SAM 30 is converted into the moreweakly bound species 38. Next the SAM 30 is treated with a displacingspecies, such as by immersion in a solution of the displacing species.As shown in FIG. 3(d) the displacing species 40 displaces the weaklybound species 38 from the substrate, and adsorbs at the surface of thesubstrate 32, resulting in a patterned SAM with a precisely definedchemical structure. The patterning is defined by relative ratios of theSNOM aperture and the SAM, and may be of a dimension commensurate withthe near field regime employed. Patterning dimensions as small as 40 nmcan be routinely achieved, and dimensions as small as 25 nm have beenproduced. It should be noted that patterned structures of largerdimensions, for example over 1 μm, can be produced using the method ofthe present invention should this be desired.

There are many possible variations upon the scheme depicted in FIG. 3which are within the scope of the invention. For example, alkyl thiolsor thiophenols may be used to form the SAM. Other types of compoundsstill which, nevertheless, form a thiolate SAM on the substrate may beused. Such compounds include dialkyl disulphides, which are of thegeneral formula R(CH₂)_(m)S—S(CH₂)_(n)R′ where R and R′ are terminalfunctional groups and m and n are each typically in the range 0 to 20.Other candidates include dialkyl sulphides, which are of the generalformula R(CH₂)_(m)S(CH₂)_(n)R′ where R and R′ are terminal functionalgroups(and may be identical), and m and n are each typically in therange 0 to 20. All of these species are candidates for use as thedisplacing species as well.

Although thiolate SAMs represent presently preferred embodiments of thepresent invention, it is possible to utilise other types of SAMs. Forexample, silane compounds might be used, eg, an alkyl silane on asilicon surface. In this instance, a possible choice of system andphotochemistry could comprise diacetylenic silanes, the diacetylenicmoieties being cross linked via photoinitiation with light from theSNOM.

It is known that monolayers of alkylsilanes on silicon can be patternedusing photolithographic methods (S. L. Brandow, M.-S. Chen, R. Aggarwal,C. S. Dulcey, J. M. Calvert and J. Dressick, Langmuir 15 (1999) 5429; S.L. Brandow, M.-S.Chen, S. J. Fertig, L. A. Chrisey, C. S. Dulcey and W.J. Dressick, Chem. Eur.J. 7 (2001) 4495; W. J. Dressick, M.-S. Chen andS. L. Brandow, J.Am. Chem. Soc. 122 (2000) 982, the contents of whichare herein incorporated by reference). In these investigations, amonolayer of chloromethylphenylsilane is formed, and then exposed to UVlight through a mask. Chloromethylphenysilanes absorb strongly at around193 nm and weakly at around 254 nm. Exposure to 193 nm light causesphotooxidation of the chloromethyl group, converting it to an aldehydefunctionality. This aldehyde functionality may be used as the site ofattachment of either organic molecules or metals. In place of the maskutilised in this prior art technique, it is possible to use excitationwith a near field light source, such as a SNOM, at around 193 nm or 254nm in order to initiate photochemistry and thereby pattern a silane SAMon a silicon surface.

Other possibilities include monolayers of carboxylic acids on, forexample, alumina surfaces, and monolayers of phosphonic acids on, forexample, indium tin oxide surfaces. It may be possible to utiliseLangmuir-Blodgett monolayer films instead of SAMs.

It is possible to utilise other photochemical reactions instead of thephotooxidation scheme depicted in FIG. 3. Diacetylenic moieties presentin the monolayer may be cross linked using the SNOM to form extendedconjugated structures. Cross linking procedures are discussed in D. N.Batchelor, S. D. Evans, T. L. Freeman, H. Ringsdorf and H. Wolf, J.Am.Chem.Soc 116 (1994) 1050-1053, and M.Cai, D. Mowery. H. Menzel and C. E.Evans, Langmuir 15 (1999) 1215-1222, the contents of which are hereinincorporated by reference. In this way, nanoscale organic circuits mightbe produced by attaching a monolayer of a suitable diacetyleniccontaining compound to a substrate and using a SNOM to trace patternscorresponding to the described circuitry on the monolayer. Along thispattern, the diacetylenic groups cross-link to form conjugated,conducting molecular wires. Diacetylenic silanes may be utilised, andthe monolayer may be formed on a silicone oxide surface. Otherphotochemical reactions might include the photocleaving of protectinggroups which exposes functionalities that are reactive with respect tosolution phase metallic species.

Another alternative photochemical scheme involves the use of a monolayercomprising molecules having photoactive end groups to which biologicalmolecules may be attached after photoactivation using light from theSNOM. Examples of such photochemical schemes can be found in E.Delamarche, G. Sundarababu, H. Biebuyck, Ch. Gerbert, H. Sigrist, H.Wolf, H. Ringsdorf, N. Xanthopoulos and H. J. Mathieu, Langmuir 12(1996) 1997-2006, and Z. Yang,W. Frey, T. Oliver and A. Chilkoti,Langmuir 16 (2000) 1751, and A. S. Blawas and W. M. Reichert,Biomaterials 19 (1998) 595, the contents of which are herebyincorporated by reference.

EXAMPLE 1

Self-assembled monolayers were prepared by immersing freshly depositedlayers of gold (30 nm) on chromium (20 nm) primed glass microscopeslides in 1 mMol dm⁻³ solutions of alkanethiols in ethanol. Thelithography experiments were carried out using a ThermoMicroscopesAurora Near-field Scanning Optical Microscope. Fused silica opticalprobes were specially manufactured by ThermoMicroscopes and coupled to afused silica fibre. The nominal internal diameter of the probes was 50nm. The optical fibre was coupled to a Coherent Innova 300C FreDfrequency-doubled argon ion laser. The fundamental wavelength at 488 nmwas doubled using a beta barium borate (BBO) crystal cut at the Brewsterangle. Features were creating by tracing the optical probe across thesample surface in a pattern controlled by the lithography software ofthe SNOM and subsequently immersing the sample in a 10 mMol dm⁻³solution of an alkyl thiol with a contrasting terminal groupfunctionality. The resulting nanometre scale patterns were imaged usinga ThermoMicroscopes Explorer Atomic Force Microscope in Lateral ForceMode.

The wavelength of 244 nm is suitable to initiate photooxidation ofthiolate species in the SAM to the relatively weakly bound sulphonatespecies. The sulphonate species are displaced by the alkyl thiol.

An HS(CH₂)₁₁OH SAM was patterned in the above described manner. Asolution of HS(CH₂)₁₁CH₃ was used to displace the sulphonate speciesproduced after SNOM patterning. The resultant patterned SAM was imagedby lateral force microscopy, which is a variant of atomic forcemicroscopy in which local variations in surface friction are measured.The image is shown in FIG. 4. Low contrast is observed in regionscorresponding to the methyl terminated SAM, because these regionsexhibit a relatively low coefficient of friction. Bright contrast isobserved in regions corresponding to the hydroxyl terminated SAM, theseregions exhibiting a higher coefficient of friction. Referring to FIG.4, two lines 50, 52 of low contrast can be seen, corresponding toadsorbed HS(CH₂)₁₁CH₃. The width of the lines 50, 52 is ca. 40 nm.

EXAMPLE 2

The method of Example 1 was utilised to pattern a SAM formed usingHS(CH₂)₁₁CO₂H. A lateral force microscopy image of the patterned SAM isshown in FIG. 5. FIG. 5(a) shows a plurality of lines 60, 62, 64, 66, 68of low contrast, corresponding to adsorbed HS(CH₂)₁₁CH₃, the lines 60,62, 64, 66, 68 being visible on a background of bright contrast,corresponding to the carboxylic acid terminated SAM. FIG. 5(b) shows aclose up view of one of the lines. The width of this line is only 25 nm.

EXAMPLE 3

The method of Example 1 was utilised to pattern a SAM formed usingHS(CH₂)₁₁CH₃. In this instance, both HS(CH₂)₁₁OH and HS(CH₂)₁₁CO₂H havebeen used successfully as the displacing species.

EXAMPLE 4

The present invention was utilised in a process for patterning goldfilms. The process is depicted schematically in FIG. 6. A gold film 70has a self-assembled monolayer (SAM) of an alkanethiol 72 formedthereon. A SNOM is disposed very close to the SAM 72 so that interactionin the near field regime can occur. The SNOM 74 is shown in FIG. 6(a),which irradiates the SAM 72 with light 76 of an appropriate wavelengthto initiate photochemistry of the type described previously, ie, thephotooxidation of the thiolate species to a relatively weakly boundsulphonate species 78. FIG. 6(b) depicts the patterned SAM 72, which nowcomprises areas of weakly bound sulphonates 78. Thereafter, thepatterned SAM 72 and gold film 70 are subjected to a chemical etch. Thechemical etch is sufficient to remove the sulphonate species 78 and etchgold underlying the sulphonate species. However, as shown in FIG. 6(c),the etch does not displace the alkanethol 72. Thus, the underlying goldfilm can be etched in a pattern defined by the SNOM. In a non-limitingexample, a gold film was covered with a SAM of hexadecanethiol. The SAMwas etched using a SNOM operating at 244 nm by way of tracing two linesin the SAM. The patterned SAM formed by this process was immersed in aferri/ferrocyanide etch solution, which removed gold underlying theoxidised regions of the SAM, eg, the portions of the gold filmunderneath the two line pattern traced on the SAM. FIGS. 7(a) and 7(b)show an atomic force microscopy image of the resulting etched film Twotrenches, corresponding to the two lines patterned by the SNOM, areclearly discernible. The width of the trenches is only 50 nm, which isequal to the diameter of the aperture in the SNOM tip 74. This exampledemonstrates that the SNOM technique of the present invention can beused in conjunction with wet etching to create metallic nanostructureswhose dimensions are effectively limited by the diameter of the aperturein the probe used for patterning. The image shown in FIG. 7 is of theetched gold substrate with the patterned SAM thereon. If desired, it isstraightforward to remove the SAM to leave the etched gold surface.Removal of the SAM can be achieved through exposure to UV light followedby washing.

It is known (Y. Xia, X-M. Zhao, E. Kim and G. M. Whitesides, Chem.Mater., 7 (1995) 2332, the contents of which are herein incorporated byreference) to apply thiols onto a metal surface using microcontactprinting, and thereafter to expose the patterns thus formed to a wetetch. This process results in removal of metal from exposed regions (ie,regions not covered with thiols). An advantage of the present inventioncompared to microcontact printing is that it generates more stablepatterns. Because the defect density in monolayers formed bymicrocontact is higher, they are more susceptible to attack by etchantsfor the gold film. In contract, the unoxidised regions of patterns formby the present invention are up to ten times more stable thancorresponding masking regions formed by microcontact printing, asignificant advantage in practical applications.

These data illustrate (i) that SNOM-patterned monolayers are very stableand (ii) that SNOM is capable of diverse applications (in other words,it is not limited to the patterning of surface chemical structuresalone).

Subsequent Functionalisation of the Patterned SAM

Judicious selection of the end groups present on the patterned monolayercan enable the selective coupling of desired molecular species to thepatterned monolayer. The molecular species might be coupled to thepatterned areas of the monolayer (ie, the lines 50, 52 in FIG. 4), or tomolecules of the monolayer itself (ie, the areas of bright contrastdepicted in FIG. 4). In principle, different molecular species might becoupled to each area. In practise, a single molecular species will becoupled to one of these areas. The identity of the compound present inthe other area where the molecular species is not intended to be presentwill be selected so that coupling does not occur. Combinations of thetype described in Examples 1 to 3 above are useful, ie, a polar endgroup such as OH or CO₂H in combination with a hydrophobic end such asmethyl. The molecular species might be coupled by adsorption of themolecular species onto a compound present on the substrate. For example,U.S. Pat. No. 5,514,501 describes various immobilisation procedures inwhich a number of biological molecules are adsorbed onto a SAM composedof carboxyl terminated thiolates. The other areas of the patterned SAMsmay be composed of thiolates that do not adsorb the biological moleculesto any great extent, for example hydroxyl (or oligo ethylene glycol)terminated thiolates.

Alternatively, covalent bonding to molecules present on the substrate ispossible. Reactive polar end groups such as OH, CO₂H and NH₂ are usefulin this regard. Alternatively still, an intermediate compound orcompounds may be coupled to a desired region of the substrate, and themolecular species coupled to the intermediate compound or compounds. Theskilled reader is directed to the wide literature that exists of varioustechniques for immobilising molecules onto surfaces. In the context ofthe present invention, what is required is that a monolayer is formed ina desired pattern having an end group which is commensurate with use ina selected immobilisation technique.

A very wide range of devices can be fabricated using the presentinvention. There are a large number of devices which might incorporate abiological molecule, such as a protein, DNA strand, RNA strand,oligonucleotide or enzyme. Some examples of such devices are describedbelow.

1. Oligonucleotide Arrays.

In one approach, a homofunctional SAM is produced on a substrate, andSNOM used to etch spots, or other desired features, onto the array usingthe techniques discussed above. A thiolate of contrasting functionalityto the originally produced SAM is formed in the spots, such as byimmersion in a suitable displacing species. A first base, coupled to aphotocleavable protecting group, is attached to the spots formed by thedisplacing species. Selected spots may be deprotected, and a furtherbase, also with a photocleavable protecting group, is attached. Adifferent sequence may be constructed at each spot by controlling arepeated sequence of deprotection and reaction steps. Such a deviceoffers the potential for building an array capable of hybridising, andhence sequencing, a single DNA molecule.

In a second approach, spots would be produced and a thiolate of acontrasting functionality formed thereon as described in the firstapproach. Thereafter, pre-synthesised oligonucleotides are attached tothe spots. The SNOM might be coupled to a microfluidic delivery systemso that rapid stepwise surface functionalisation can be performed.

2. Protein Arrays

Protein arrays can be created in a similar way to the oligonucleotidearrays to discussed above, except that attachment to the spots involvesthe use of covalent chemistry to attach proteins. The covalentattachment chemistry is well established in the literature. Anattraction of coupling the known protein attachment chemistry with thepatterning method of the present invention is that it becomes possibleto produce nanoscale analogues of processes which are well establishedat larger scales.

3. Photo-Activated Coupling Agents for Biological Arrays

In this scheme, photocleavable protecting groups are covalently attachedto an unpatterned SAM. Treatment by the SNOM results in deprotection ofthe protecting groups to create ‘active’ features. Molecules such asproteins and oligonucleotides may then bind to the active features. Sucha process might be carried out in liquid, holding out the possibilitythat rapid sequential attachment steps might be performed.

1-24. (Canceled)
 25. A method of patterning a monolayer comprising thesteps of: providing a self-assembled monolayer of a compound on asubstrate; positioning a near field light source in relation to themonolayer so that light from the light source irradiates the monolayerin the near field regime, the wavelength of the light being suitable tointeract with molecules in the monolayer and thereby initiate aphotochemical reaction; and patterning the monolayer by causing relativemovement of the monolayer and the near field light source, the relativemovement corresponding to a desired pattern.
 26. A method according toclaim 25, wherein the near field light source comprises a scanning nearfield microscope.
 27. A method according to claim 25, wherein themolecules in the monolayer which absorb light from the light source areconverted by the photochemical reaction into a weakly bound specieswhich is less strongly bound to the substrate than the molecules whichoriginally comprise the monolayer.
 28. A method according to claim 27,further comprising displacing the weakly bound species from thesubstrate with a displacing species.
 29. A method according to claim 25,wherein the photochemical reaction is a photooxidation reaction.
 30. Amethod according to claim 25, wherein the molecules in theself-assembled monolayer comprise thiolates.
 31. A method according toclaim 30, wherein the self-assembled monolayer is formed from thiols.32. A method according to claim 31, wherein said thiols comprise alkylthiols.
 33. A method according to claim 31, wherein said thiolscorrespond to the formula HS(CH₂)_(n)X, where X is an end group.
 34. Amethod according to claim 33, wherein group X is selected from the groupconsisting of CH₃, CO₂H and OH.
 35. A method according to claim 29,wherein the photooxidation reaction oxidizes a thiolate moiety adsorbedon the substrate to a sulphonate moiety.
 36. A method according to claim28, wherein the displacing species forms a thiolate compound on thesubstrate.
 37. A method according to claim 25, wherein the substratecomprises gold or silver.
 38. A method according to claim 25, whereinthe light source provides near UV light.
 39. A method according to claim25, wherein the patterning comprises features having at least onedimension in a direction parallel to the substrate which is less than100 nm.
 40. A method according to claim 39, wherein the patterningcomprises features having at least one dimension in a direction parallelto the substrate which is less than 50 nm.
 41. A method of selectivelycoupling a molecular species to a surface, comprising the steps of:patterning a monolayer using a method according to claim 25 to provideat least one feature; and coupling the molecular species to a compoundpresent on the substrate.
 42. A method according to claim 41, whereinthe molecular species is coupled to the monolayer.
 43. A methodaccording to claim 41, wherein the molecular species is coupled to theat least one feature.
 44. A method according to claim 43, wherein in themethod used to pattern the monolayer the molecules in the monolayerwhich absorb light from the light source are converted by thephotochemical reaction into a weakly bound species which is lessstrongly bound to the substrate than the molecules which originallycomprise the monolayer; the method used to pattern the monolayer furthercomprises displacing the weakly bound species from the substrate with adisplacing species, and the molecular species is coupled to thedisplacing species.
 45. A method according to claim 41, wherein at leastone intermediate compound is coupled to a portion of the patternedmonolayer.
 46. A method according to claim 41, wherein the couplingcomprises adsorption of the molecular species onto the compound.
 47. Amethod according to claim 41, wherein the coupling comprises covalentlybonding the molecular species to the compound.
 48. A method according toclaim 41, wherein the molecular species is a biological molecule.
 49. Amethod according to claim 48, wherein the biological molecule is aprotein, a DNA strand, an RNA strand, an oligonucleotide or an enzyme.